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Bacterial community analysis of activated sludge: an evaluation of four commonly used DNA extraction methods

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Abstract

The effectiveness of three commercially available direct DNA isolation kits (Mobio, Fast, Qiagen) and one published direct DNA extraction protocol (Bead) for extracting bacterial DNA from different types of activated sludge was investigated and mutually compared. The DNA quantity and purity were determined using real-time PCR targeting the bacterial 16S rDNA gene. Microbial community fingerprints were assessed by automated ribosomal intergenic spacer analysis. The resulting community profiles were analyzed with canonical correspondence analysis. Our results clearly demonstrate that direct DNA extraction methods can significantly influence the DNA quantity, purity, and observed community patterns of microbiota in activated sludge. Fast and Mobio generated high amounts of good quality DNA compared to Bead and Qiagen. Mobio also resulted in the detection of the highest number of species while Fast scored the best in discriminating between the community patterns of different activated sludge types. With respect to the characterization of community profiles, our analyses demonstrated a strong sludge type dependent variability among methods. Taking into account our results, we recommend Fast as the most suitable DNA extraction method for activated sludge samples used for bacterial community studies.

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Acknowledgments

This work was supported by the IDO/06/008 project, Katholieke Universiteit Leuven Research Fund. Louise Vanysacker acknowledges the Fund for scientific research Flanders for a PhD scholarship. We thank M. Feyeaerts (Enprotech, Rotselaar, Belgium) and X. Yan (Waterleau, Wespelaar, Belgium) for kindly providing the activated sludge samples.

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Authors and Affiliations

  1. Centre for Surface Chemistry and Catalysis, Katholieke Universiteit Leuven, Kasteelpark Arenberg 23, P.O. Box 2461, 3001, Heverlee, Belgium

    Louise Vanysacker & Ivo Vankelecom

  2. Laboratory of Aquatic Ecology and Evolutionary Biology, Katholieke Universiteit Leuven, Charles Deberiotstraat 32, 3000, Leuven, Belgium

    Steven A. J. Declerck, Luc De Meester & Priscilla Declerck

  3. Laboratory of Animal Diversity and Systematics, Katholieke Universiteit Leuven, Charles Deberiotstraat 32, 3000, Leuven, Belgium

    Bart Hellemans

  4. Department of Aquatic Ecology, Netherlands Institute of Ecology (NIOO-KNAW), Rijksstraatweg 6, 3631 AC, Nieuwersluis, The Netherlands

    Steven A. J. Declerck

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  1. Louise Vanysacker
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  2. Steven A. J. Declerck
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  3. Bart Hellemans
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Correspondence to Louise Vanysacker.

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Vanysacker, L., Declerck, S.A.J., Hellemans, B. et al. Bacterial community analysis of activated sludge: an evaluation of four commonly used DNA extraction methods. Appl Microbiol Biotechnol 88, 299–307 (2010). https://doi.org/10.1007/s00253-010-2770-5

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